The overall objective of this proposal is to understand the potential intracellular mediators involved in the mechanism of action of immune cell factors on osteoblastic cells. With the use of recombinant material it has been recently shown that the monocyte products, interleukin-1 and tumor necrosis factor alpha, as well as the lymphocye product, lymphotoxin, have direct stimulatory effects on bone resorption. Another lymphocyte product, interferon gamma, appears to be directly involved in the inhibitory regulation of the resorption induced by the other factors. Osteoblasts appear to be directly involved in the regulation of resorption by these and other resorptive agents. the effects of these factors on osteoblastic cell calcium will be studied using the calcium-sensitive fluorescent dye, Fura-2. Cyclic AMP metabolism will be studied by radioimmunoassay and by direct measurement of adenylate cyclase activity. Endogenous synthesis of prostaglandin E2 will be studied by a specific radioimmunoassay and by incorporation and release of 14C arachidonic acid. The interrelationships among these putative mediators will also be studied with the use of inhibitors of cyclooxygenase, agents such as forskolin and phosphodiesterase inhibitors, which increase cyclic AMP levels, and with the use of agents such as calcium inophores and antagonists which influence cytosolic calcium. The immune cell factors will be studied alone and in combination with each other and with the local hormone, prostaglandin E2, and with the systemic hormones, parathyroid hormone and 1,25(OH2)D3. Preparations of normal rat osteoblasts obtained by sequential collagenase digestions of newborn calvaria as well as osteosarcomal clones and subclones representing various stages of differentiation will be studied. Since these immune cell factors are released during inflammatory reactions, an understanding of their mechanism of action should aid in the development of therapies directed toward the control of bone loss that occurs as a host response in oral infectious diseases.